The mechanisms of intestinal secretion will be explored in this application by using large organic anions to inhibit active, carrier mediated transport processes in intestinal epithelial cells and by study of these transport processes in vesicles created from isolated brush border and basolateral cell membranes of these cells. Inhibitors of anion transport in red blood cells, such as the disulfonic stilbenes (SITS or DIDS) and drugs of the class which include furosemide, will be studied in vitro with the Ussing chamber technique or with the influx technique of Schultz. Their ability to inhibit or stimulate net absorption or secretion of Na and Cl will be determined with radioactive tracer studies, and the results compared to previous studies in this laboratory with aspirin and phenolphthalein (also large organic anions) which appear to effect coupled NaCl transport processes in intestine. In addition, clinically important diarrheal (bile salts and long chain fatty acids) and antidiarrheal (gallic acid, pectins, tannins, heavy metals-Al and Bi, and narcotic derivatives) compounds will be similarly investigated to see if their effects are similar to the RBC transport inhibitors. Vesicles will be made from luminal and contraluminal cell membranes from both villous and crypt enterocytes after purification of those membranes with standard techniques. The location and characterization of coupled NaCl transport process will be determined in these membranes by demonstrating the ability of Cl (and not other anions) to promote Na transport and Na (and not other cations) to promote Cl transport under both gradient and nongradient conditions, with saturation kinetics, and in experiments where transmembrane potentials cannot account for the stimulated transport. The organic anions will be tested for their ability to inhibit these transport processes. If successful, these experiments will confirm which of the current models for intestinal secretory processes is more correct, and will establish a paradigm which explains the action of diarrheal and antidiarrheal drugs. This investigation should advance our basic understanding of intestinal electrolyte transport processes and should have direct practical application to an understanding of the mechanisms and treatment of diarrhea.